The nucleotide excision repair (NER) pathway is responsible for the recognition and repair of a diverse array of helix distorting DNA lesions and is therefore crucial in the maintenance of genomic stability. Studies have revealed that NER can be further subdivided into the fast transcription coupled repair (TCR) pathway, which preferentially repairs transcribed genes, and the slower global genome repair (GGR) pathway, which repairs the remainder of the genome. With the discovery that sequence-specific triplex forming oligonucleotides (TFOs) could bind as a third strand to duplex DNA to create a non-canonical helical structure, a method of introducing helix distorting lesions to monitor repair at specific sites in the genome became possible. The long-term objective of this proposal is to use the technology of producing site-specific helical alterations to understand the relative contributions of the TCR and GGR pathways in the repair of altered DNA structures. The specific aims of this proposal are: 1. To compare the roles of the TCR pathway versus the GGR pathway in the repair of unusual DNA structures. 2. To examine the repair and recognition of different helix distorting molecules. Experiments designed to monitor recognition and repair of unusual DNA structures will be performed using combinations of a variety of site-specific DNA binding molecules, cell-free extracts derived from human repair deficient cell lines, and specific antibodies to DNA repair factors and purified proteins. Specifically, the roles of the GGR protein, XPC, and the TCR protein, CSB, in recognition and repair of altered DNA structures will be examined and compared to the role of the NER protein, XPA.